Breast cancer should be viewed as an organ system in which growth and progression are governed by complex and reciprocal interactions between tumor cells and surrounding stromal elements. Fibroblasts, which comprise a predominant stromal cell type, maintain tissue homeostasis in normal breast but promote tumor progression in breast cancer. Carcinoma-associated fibroblasts (CAF) distinguish themselves from normal mammary fibroblasts (NMF) by morphology, gene expression and secreted factors. Our lab has shown that expression of the cell surface proteoglycan syndecan 1 (Sdc1) in CAF is induced in the majority of breast carcinomas and that Sdc1 stimulates breast carcinoma proliferation. Because one of the main functions of fibroblasts is the assembly of an extracellular matrix (ECM), we have begun to examine whether Sdc1 expression affects ECM synthesis. Our preliminary data indicate that Sdc1 expression in CAF influences the architecture, or fine structure, of the ECM scaffold. It is the goal of this proposal to understand in detail how Sdc1 regulates ECM assembly in breast carcinomas and what consequences Sdc1-dependent ECM alterations might have on carcinoma behavior. Based on our preliminary observations, we state the following hypothesis: The aberrant expression of Sdc1 by breast carcinoma stromal fibroblasts leads to an altered ECM architecture, which is permissive to breast carcinoma cell invasion. We posit that this altered ECM architecture contributes to invasion events early and late during the natural history of the disease and shortens patient survival. To test this hypothesis, we propose the following specific aims: Aim 1: Examine the role of Sdc1 and ECM architecture in breast carcinoma invasion. The ECM architecture will be carefully analyzed in human breast carcinoma samples. Using tissue microarrays, we will determine whether ECM architectural features predict patient prognosis. Innovative ex vivo invasion assays will inform us whether Sdc1 and/or the ECM architecture regulate invasion. Lastly, the involvement of Sdc1 in determining the ECM architecture will be examined with Sdc1-deficient animals. Aim 2: Analyze the role of Sdc1 and ECM architecture in the progression from ductal carcinoma in situ (DCIS) to invasive carcinoma. By applying novel ECM imaging tools to human samples and to a DCIS animal model, we will determine whether stromal Sdc1 expression creates an invasion-permissive ECM that facilitates progression from DCIS to invasive carcinoma. Aim 3 Decipher the molecular mechanisms responsible for the formation of an invasion-permissive ECM. The involvement of specific Sdc1 molecular domains in regulating ECM assembly will be analyzed in vitro with domain deletion and substitution experiments. The cooperative role of integrin cell adhesion receptors will be investigated with loss of function and gain of function experiments. Together, these aims will significantly advance our knowledge about the regulation of ECM production in breast cancer. A mechanistic understanding of ECM assembly is key to the design of novel therapeutic agents that are aimed at "normalizing" the ECM and thus revert the tumor microenvironment from invasion-permissive to invasion-restrictive. PUBLIC HEALTH RELEVANCE: With approximately 200,000 newly diagnosed cases in the US per year, breast cancer is and remains a major health issue. Even with the advanced treatment options available, 40,000 women die annually. Our project aims at understanding how the architecture of the connective tissue scaffold in breast cancer is regulated and how it affects cancer invasion and spread. The knowledge generated with this work may lead to novel therapies that target the structure of this scaffold and prevent the escape of cancer cells.